| Summary: | This work aims to determine the impact of dietary supplementation of polysaccharide, extracted from brown seaweeds <i>Sargassum dentifolium</i> on growth indices, feed utilization, biochemical compositions, microbial abundance, expressions of growth and immunity-related genes, and stress genes of the Pacific Whiteleg shrimp <i>Litopenaeus vannamei</i>. A total of 360 post-larvae of <i>L. vannamei</i> were randomly distributed into a 12-glass aquarium (40 L of each) at a stocking density of 30 shrimp with an initial weight of (0.0017 ± 0.001 g). During the 90-day experiment trial, all shrimp larvae were fed their respective diets at 10% of total body weight, three times a day. Three experimental diets were prepared with different seaweed polysaccharide (SWP) levels. The basal control diet had no polysaccharide level (SWP<sub>0</sub>), while SWP<sub>1</sub>, SWP<sub>2</sub>, and SWP<sub>3</sub> contained polysaccharides at concentrations of 1, 2, and 3 g kg<sup>−1</sup> diet, respectively. Diets supplemented with polysaccharide levels showed significant improvements in weight gain and survival rate, compared to the control diet. Whole-body biochemical composition and the microbial abundance (the total count of heterotrophic bacteria and <i>Vibrio</i> spp.) of <i>L. vannamei</i> showed significant differences among polysaccharide-treated diets compared to the control. At the end of the feeding experiment, the dietary supplementation of polysaccharide levels enhanced the expression of growth-related genes (Insulin-like growth factors (<i>IGF-I, IGF-II</i>), immune-related genes (<i>β</i> -Glucan-binding protein (<i>β-Bgp</i>), Prophenoloxidase (<i>ProPO</i>), Lysozyme (<i>Lys</i>), and <i>Crustin</i>), and stress genes (Superoxide dismutase (<i>SOD</i>) and Glutathione peroxidase (<i>GPx</i>) in the muscle tissue of <i>L. vannamei</i>. However, the current study concluded that the inclusion rate of 2 g kg<sup>–1</sup> of polysaccharide as a dietary additive administration enhanced both weight gain and survival rate of <i>L. vannamei</i>, while the incorporation level of 3 g kg<sup>–1</sup> reduces the abundance of pathogenic microbes and enhances the growth-, immunity- and stress-related gene expressions of <i>L. vannamei</i>.
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